Wren B W, Clayton C L, Mullany P P, Tabaqchali S
Department of Medical Microbiology, St Bartholomews Hospital Medical College, West Smithfield, London, England.
FEBS Lett. 1987 Dec 10;225(1-2):82-6. doi: 10.1016/0014-5793(87)81135-3.
Clostridium difficile toxin A was purified to homogeneity and was used to raise monospecific antiserum in rabbits. A gene bank of C. difficile DNA in Escherichia coli was constructed by cloning Sau3A-cleaved clostridial DNA fragments into the bacteriophage vector lambda EMBL3. Out of 4500 plaques screened with antitoxin A, 9 clones were positively identified. One of these clones lambda tA5 expressed a 235 kDa protein which exhibited a cytotonic effect on Chinese hamster ovary cells, and had the ability to haemagglutinate rabbit erythrocytes, both properties characteristic of toxin A. The size of the lambda tA5 insert DNA was 14.3 kb.
艰难梭菌毒素A被纯化至同质，并用于在兔中产生单特异性抗血清。通过将经Sau3A切割的梭菌DNA片段克隆到噬菌体载体λEMBL3中，构建了艰难梭菌DNA在大肠杆菌中的基因文库。在用抗毒素A筛选的4500个噬菌斑中，有9个克隆被阳性鉴定。其中一个克隆λtA5表达一种235 kDa的蛋白质，该蛋白质对中国仓鼠卵巢细胞表现出致细胞紧张效应，并具有凝集兔红细胞的能力，这两种特性均为毒素A所特有。λtA5插入DNA的大小为14.3 kb。
